Tumors are potentially immunogenic. However, they fail to spontaneously induce immune responses capable of^ rejecting tumors. A major reason for this is that the tumor microenvironment lacks adequate innate immune activation required to initiate strong adaptive antitumor immunity. Plasmacytoid dendritic cells (pDCs) are highly specialized in that they sense microbial nucleic acids via intracellular Toll-like receptors. During viral infection, pDCs accumulate in infected tissues and are activated by viral nucleic acids to produce large amounts of type I interferons (IFNs) and generate protective immunity against the virus by activating myeloid dendritic cells, T cells, and natural killer cells. Tumors also contain pDCs but do not provide molecular signals to activate pDCs, although tumors contain self-DNA released in the extracellular environment at high concentrations as a result of increaised turnover of tumor cells. pDCs, though activated by viral nucleic acids, clearly are normally not able to sense tumor-derived DNA and are thus unable to initiate strong innate immune responses. We recently found that pDCs can, in fact, sense and respond to self-DNA when combined with an endogenous peptide called LL37. LL37 can bind to self-DNA fragments released by dying cells to form aggregates and condensed structures that are delivered to and retained within early endosomes of pDCs.. In these intracellular compartments, LL37/self-DNA can interact with Toll like receptor 9 to trigger robust type I IFN production similariy to viral DNA. Because tumors release large amounts of self-DNA and contain pDCs but do not express LL37, our hypothesis for the proposed study described herein is that exogenous LL37 can be used to target tumor-derived self-DNA and convert it into a 'danger signal that triggers pDC activation and type I IFN production at the tumor site in patients with melanoma. This then induces T-cell-mediated immunity against melanoma by using the same mechanism by which anti-viral-immune responses are induced. Therefore, we will pursue the followinig: Specific Aim 1: Determine the mechanism of anti-tumor immune responses induced by intratumoral LL37 injection in mouse tumor models; Specific Aim 2: Evaluate anti-tumor immune responses and clinical efficacy of intratumoral LL37 injection in patients with melanoma; Specific Aim 3: Improve anti-tumor immune responses and efficacy of intratumoral LL37 injection in mouse tumor models. These studies may lead to principles in cancer immunotherapy that may be vyidely applicable to other cancers.